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Evaluation of rapid DNA extraction methods to better enable point-of-use environmental DNA detection

Melisa E. Kozaczek·Stephen Frank Spear·Tyler J. Untiedt·Paul Albosta·Caden Jungbluth·Jared J. Homola·Elliott P. Barnhart·Christopher M. Merkes
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Abstract

Recent developments in molecular testing have created the opportunity for biologists and managers to detect environmental DNA (eDNA) of target species rapidly and without the requirement of a laboratory. These point-of-use protocols may be especially useful for early detection and rapid response for invasive species or surveillance for at-risk native species, where timely management decisions are critical. Point-of-use eDNA protocols also facilitate wider and less expensive implementation of eDNA methods. One of the key components to an effective point-of-use protocol is a rapid DNA extraction method. Several rapid extraction protocols are suitable for implementation in the field, but information regarding their relative effectiveness is lacking. We evaluated extraction efficiency of four DNA rapid extraction protocols using filters spiked with primary cultured grass carp ( Ctenopharyngodon idella ) gill cells. The extraction methods included two syringe-based column extractions, a lysis and extraction solution, and a divalent cation chelation resin (Chelex) extraction protocol alongside a laboratory-based control kit. We estimated DNA yield using a newly designed quantitative polymerase chain reaction (qPCR) assay targeting the grass carp nuclear genome. We evaluated two additional factors, filter type (mixed cellulose ester [MCE] and polyethersulfone [PES]) and background eDNA source (aquaculture or river). The lysis and extraction solution and Chelex extraction both had the highest overall yield, with MCE filters further increasing Chelex yield while the enzyme extraction yield was dependent on interaction with both filter and eDNA source. Our results indicate that rapid extraction protocols, such as solutions with short heating steps, are effective for DNA isolation and help to increase the overall accessibility of eDNA analyses.

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Melisa E. Kozaczek, Stephen Frank Spear, Tyler J. Untiedt, Paul Albosta, Caden Jungbluth, Jared J. Homola, Elliott P. Barnhart, Christopher M. Merkes. 2025-07-15. Evaluation of rapid DNA extraction methods to better enable point-of-use environmental DNA detection. https://doi.org/10.1002/edn3.70159

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